Liu Yi-Chang, Fujimori Danica Galonić, Weissman Jonathan S
Chemistry and Chemical Biology Graduate Program, University of California, San Francisco, CA 94158; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158; Howard Hughes Medical Institute, University of California, San Francisco, CA 94158;
Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158; Department of Pharmaceutical Chemistry; University of California, San Francisco, CA 94158;
Proc Natl Acad Sci U S A. 2016 Jul 12;113(28):E4015-24. doi: 10.1073/pnas.1608795113. Epub 2016 Jun 28.
Our understanding of how the endoplasmic reticulum (ER)-associated protein degradation (ERAD) machinery efficiently targets terminally misfolded proteins while avoiding the misidentification of nascent polypeptides and correctly folded proteins is limited. For luminal N-glycoproteins, demannosylation of their N-glycan to expose a terminal α1,6-linked mannose is necessary for their degradation via ERAD, but whether this modification is specific to misfolded proteins is unknown. Here we report that the complex of the mannosidase Htm1p and the protein disulfide isomerase Pdi1p (Htm1p-Pdi1p) acts as a folding-sensitive mannosidase for catalyzing this first committed step in Saccharomyces cerevisiae We reconstitute this step in vitro with Htm1p-Pdi1p and model glycoprotein substrates whose structural states we can manipulate. We find that Htm1p-Pdi1p is a glycoprotein-specific mannosidase that preferentially targets nonnative glycoproteins trapped in partially structured states. As such, Htm1p-Pdi1p is suited to act as a licensing factor that monitors folding in the ER lumen and preferentially commits glycoproteins trapped in partially structured states for degradation.
我们对于内质网（ER）相关蛋白降解（ERAD）机制如何有效靶向终末错误折叠蛋白，同时避免新生多肽和正确折叠蛋白被误识别的理解仍然有限。对于内质网腔中的N-糖蛋白而言，其N-聚糖的去甘露糖基化以暴露出末端α1,6-连接的甘露糖是通过ERAD进行降解所必需的，但这种修饰是否特定于错误折叠蛋白尚不清楚。在此，我们报道甘露糖苷酶Htm1p和蛋白二硫键异构酶Pdi1p（Htm1p-Pdi1p）的复合物在酿酒酵母中作为一种对折叠敏感的甘露糖苷酶，催化这一关键的第一步。我们用Htm1p-Pdi1p和结构状态可操控的模型糖蛋白底物在体外重建了这一步骤。我们发现Htm1p-Pdi1p是一种糖蛋白特异性甘露糖苷酶，优先靶向被困于部分结构化状态的非天然糖蛋白。因此，Htm1p-Pdi1p适合作为一种许可因子，监测内质网腔中的折叠情况，并优先将被困于部分结构化状态的糖蛋白提交进行降解。
